1. Field of the Invention
The present invention relates generally to directed beam irradiation devices and specifically to an improved method and apparatus for mounting and maintaining specimens for observation and characterization in a scanning electron microscope.
2. Discussion of the Prior Art
The scanning electron microscope is a powerful instrument which permits observation and characterization on a micrometer or submicrometer scale of heterogeneous organic and inorganic materials. Limited microscopic techniques are available for examining volatile i.e., "wet" samples and biological specimens, where hours of tedious preparation usually is required, resulting in a non-living and disrupted state. Either a relatively low resolution and/or limited depth of focus optical microscope or a specialized high voltage transmission electron microscope equipped with gas reaction cells must be used. The SEM with its great advantage of depth of focus, has only limited capabilities for maintaining a vapor pressure of even a few Torr.
In a SEM, an electron beam is scanned across the specimen to be observed and then electrons reflected or generated from the specimen are detected and the signal generated by the detector is processed to form the enlarged image. In order to properly scan a specimen, an extremely low pressure on the order of 10.sup.-4 TORR is necessary in the specimen chamber of the electron microscope. This extremely low pressure is dramatically less than the vapor pressure of water and a number of other liquids comprising structures of interest. Therefore, in attempting to examine solids, such as hydrated cements or biological samples, inevitably, upon exposure to the high vacuum of the electron microscope, fluid in the solid evaporates rupturing and otherwise destroying the structural arrangement of the subject under investigation. A conventional system in which tissue samples are sectioned while frozen in order to form a thin foil still lose most of their water content immediately upon exposure to the high vacuum rendering them impossible to study. Dry preparation techniques for thin foil specimens are tedious and complicated, often resulting in disruption of and artifact formation in the specimen.
There have been numerous attempts, by freezing or other methods, to slow down the specimen destruction usually meeting with little success. One such method is disclosed in U.S. Pat. No. 4,071,766 to Kalman et al which utilizes a pressure-balancing buffer system to provide a pressure in the vicinity of the specimen which is greater than the vapor pressure of the liquid in the specimen in order to prevent rapid bursting or deterioration. This system utilizes a separate tank containing a liquid or gas which is connected to the specimen mounting chamber and maintains the desired pressure and moist atmosphere to aid in the preservation of the specimen.
The difficulty with such pressure balancing systems is the requirement of an auxiliary pump connected to the electron microscope in order to maintain the high vacuum necessary for the operation of the microscope. Thus, after mounting of the specimen in the microscope, the various pumps are energized and as the microscope nears its operational high vacuum, the balancing system continually evolves a vapor in the chamber in order to attempt to prevent destruction of the specimen. Therefore, the buffer system is generating a vapor and the vacuum pumps are pulling that vapor away in an attempt to reach operational pressure. The buffer system and the vacuum pumps are working at cross purposes and a substantial amount of time is wasted in attempting to reach a stabilized condition in order to begin the electron microscopy. This constant evolution of vapor and then pumping away of the evolved vapor insures that the electron beam must penetrate a substantial thickness of vapor surrounding the specimen in order to accomplish its scanning function. The thickness of vapor penetrated affects the quality of the resultant micrograph and, even though the pressure buffer attempts to maintain the pressure at or about the vapor pressure of the liquid contained in the specimen, deterioration of the specimen is likely.